The goal of this project is to define the physiological significance of an apparently novel protein that stimulates growth of kidney epithelial cells. We previously identified a protein that appears when nontransformed monkey renal epithelial cells (BSC-1 line) are exposed for 3-5 minutes to a reduced concentration of sodium in the culture medium; it is called the low sodium growth factor (LSGF). This protein has an apparent molecular weight about 6,500, stimulates growth of both monkey and canine kidney epithelial cells in culture but not fibroblasts, resists denaturation by dithiothreitol, appears to differ from known growth factors of similar size with respect to biological activity and amino acid composition, and resides in the extracellular matrix from which it is released when the sodium concentration of the medium is reduced. The amino terminus of the LSGF protein is blocked. A protocol to purify LSGF by reversed-phase (RP) HPLC was developed during the 03 grant year. This revised application describes a new biologically active isoform of LSGF detected by RP HPLC that has a different amino acid composition than the previously described protein. In addition, a rabbit antiserum against LSGF has been prepared that recognizes both LSGF isoforms of ELISA, and also neutralizes the growth-promoting activity of each. To define the structure of LSGF and mechanisms by which it exerts its physiological effects on renal cells in vitro and in vivo, new reagents are required that will be used to achieve the following specific aims: 1. partially sequence the LSGF protein; 2. use oligonucleotide probes or immunoscreening to clone the gene that encodes the protein, and study regulation of its expression during cell growth; 3. develop LSGF immunoassays to study the physiology the protein; 4. determine if LSGF alters nutrient and ion transport in renal epithelial cells. Achieving these aims could define the role of this autocrine growth factor in renal physiology and pathophysiology. Identifying mechanisms by which the LSGF acts may be an important step towards our long-term goal of formulating a therapeutic solution containing specific components such as nutrients and growth factors that could speed regeneration of the kidney after injury.